Week 1

This first week has been very exciting and and informative. After being bombarded with journal articles, slide shows and lectures on various aspects of my project, I began to understand the gist of it.

Dr. Zuber's lab has for some time been studying bacterial responses to environmental stress. Within the last few years they have identified a protein they lovingly call Spx, which the soil bacteria Bacillus subtilis produces upon oxidative stress. Spx interacts with the alpha subunit of RNA polymerase and hinders transcription of certain genes (mainly those associated with growth, competence and sporulation) and promotes genes that encode proteins which have roles in thiol homeostasis. Spx is normally kept at low levels via the ATP-dependent protease ClpXP. Spx is ushered to the protease complex by the accssory protein YjbH.

My job this summer will be to study the effects of different mutations upon Spx and explore how these mutations effect the regulation of this protein within the cell (i.e. The interaction between Spx and ClpXP). I will also be exploring other cellular targets of YjbH and the comparing its effect on other proteins with its role in Spx regulation.

This week I have grown cells containing a G42R Spx mutation which is believed to interupt the Spx/RNAP interaction. I have been working on isolating and purifying the Spx protein from these bacteria. After using a French press to lyse the B. subtilis I used His-tag Ni-NTA column chromatography to isolate my desired protein. These samples were stored in 4 degrees C until next week when I will use SDS-PAGE to view my protein.